Multiphoton microscopy enables deep imaging with less overall photobleaching than with wide-field or confocal microscopy, and it has now become the primary fluorescence imaging technique for a variety of biological samples and systems. For these difficult imaging tasks, quantum dots (QDs), mostly those based on nanoscale semiconductors, are considered as promising fluorescent agents for in vivo multiphoton imaging. However, a significant shortcoming for some of the best-performing QDs is their containing heavy metals such as cadmium or lead, which pose potential health and environment risks and limit their clinical uses. Nanosized pure carbon particles may be surface-passivated to exhibit strong photoluminescence in the visible and near- infrared regions, which may be considered effectively as carbon quantum dots. Recent results suggest that carbon quantum dots have very large multiphoton cross-sections, offering the potential to serve as benign and competitive alternatives to traditional semiconductor QDs-based multiphoton imaging agents. Among the known and expected significant advantages for carbon quantum dots include carbon being an abundant and nontoxic element, the physicochemical and photochemical stability of the materials, and the likely low costs in the eventual large-scale production of the materials as imaging agents. In this project the research team will pursue two specific aims: (1) to evaluate, improve, and optimize the multiphoton luminescence properties of carbon quantum dots through the understanding of parameters that affect the performance of the dots as multiphoton imaging agents; and (2) to begin to address the key issues important to the commercialization of the technology, including the selection and evaluation of various sources of carbon particles, the standardization of the carbon dots configuration, and the bioconjugation of carbon dots in preparation for their tests in imaging systems. The ultimate goal of the project is to quickly develop the technology toward maturation and commercialization. [unreadable] [unreadable] [unreadable]